Characterization of Tailoring Steps of Nargenicin A1 Biosynthesis Reveals a Novel Analogue with Anticancer Activities

Abstract

Nargenicin A1(1) is an antibacterial macrolide with effective activity against various Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus. Due to the promising properties of this compound in inhibiting cell proliferation, immunomodulation, and the cell protective effect, there has been significant interest in this molecule. Recently, the biosynthetic gene cluster (BGC) of 1 was reported from Nocardia argentinesis and Nocardia arthritidis. In addition, two crucial enzymes involved in the formation of the core decalin moiety and postmodification of the decalin moiety by an ether bridge were characterized. This study reports on the BGC of 1 from Nocardia sp. CS682. In addition, the direct capture and heterologous expression of nar BGC from Nocardia sp. CS682 in Streptomyces venezuelae led to the production of 1. Further metabolic profiling of wild type, Nocardia sp. CS682 in optimized media (DD media) resulted in the isolation of two acetylated derivatives, 18-O-acetyl-nodusmicin and 18-O-acetyl-nargenicin. The post-PKS modification pathway in biosynthesis of 1 was also deciphered by identifying intermediates and/or in vitro enzymatic reactions of NgnP1, NgnM, and NgnO3. Different novel analogues of 1, such as compound 6, compound 7, 23-demethyl 8,13-deoxy-nodusmicin (8), 23-demethyl 8,13-deoxynargenicin (9), 8,13-deoxynodusmicin (10), and 8,13-deoxynargenicin (11), were also characterized, which extended our understanding of key post-PKS modification steps during the biosynthesis of 1. In addition, the antimicrobial and anticancer activities of selected analogues were also evaluated, whereas compound 9 was shown to exhibit potent antitumor activity by induction of G2/M cell cycle arrest, apoptosis, and autophagy.